Surgical treatment of musculoskeletal diseases relies more and more on the long-term implantation of foreign materials such as bone substitutes, endoprosthesis, degradable scaffolds and plastic components. Since the immune system is not adapted well to fight bacterial infection associated with these foreign materials, septic complications are a growing concern for the orthopedic community (Antoci et al. Clin Orthop Relat Res. 2007; 461:81-87 Antoci et al. J Orthop Res. 2007; 25:858-866). Due to the low metabolic rate of bone tissue and the formation of bacterial biofilm, it is difficult to reach the required local concentration of antibiotics whether it is applied systemically or on the spot during surgery (Costerton et al. Science. 1999; 284:1318-1322, Ketonis et al. Antimicrob Agents Chemother. 2011; 55:487-494., Stoodley et al. Annu Rev Microbiol. 2002; 56:187-209). In the general practice, local treatment is typically applied to support systemic antibiotics, the most frequently used drugs include amoxicillin, cephalexin, gentamicin, sulfamethoxazole, ciprofloxacin and vancomycin, applied in cement (Buchholz et al. Clin Orthop Relat Res. 1984:96-108, Trippel et al. J Bone Joint Surg Am. 1986; 68:1297-1302, Wininger et al. Antimicrob Agents Chemother. 1996; 40:2675-2679.), beads and impregnated bone (Barckman et al. J Biomed Mater Res B Appl Biomater. 2013, Buttaro et al. Hip Int. 2010; 20:535-541, Ketonis et al. Clin Orthop Relat Res. 2010; 468:2113-2121, Ketonis et al. Bone. 2011; 48:631-638, Melichercik et al. Folia Microbiol (Praha). 2012; 57:459-462, Winkler et al. Int J Med Sci. 2009; 6:247-252). In addition, off-label use of these antibiotics mixed by hand with the carrier bone substitute is often performed when the required antibiotic-carrier combination is not available off-the-shelf (Mathijssen et al. BMC Musculoskelet Disord. 2012; 13:44). Whether the applied dose and the release kinetics of such mixtures is optimal or at least adequate for the intended purpose is unknown, but it is still the best way a surgeon can deal with these challenging situations.
The therapeutic goal of local antibiotic use in combination with endoprostheses can be categorized into three distinct case types: 1, prevention of early infection at a primary prosthesis implantation procedure, 2, inhibiting infection at aseptic prosthesis revisions where the probability of an already ongoing low-grade infection is high and 3, treatment of massive infections at septic revisions (Gehrke et al. Hip Int. 2012; 22 Suppl 8:S40-45, Klatte et al. J Arthroplasty. 2013). These cases pose differing challenges for the antibiotics summarized in Table 1.
TABLE 1Technical requirements against a local antibiotic formulation in the3 main categories of orthopedic use in endoprosthesis surgery.Required lengthTypical localProbability ofof local antibioticantibioticMedical purposeinfectionAntibiogramtreatmentformulationPrimaryPrevention of0.5-2%Not available1-2 days or untilAntibiotic boneimplantationinfection arisingthe surgical sitecement, off-the-fromis open throughshelfcontamination atdrainagesurgery or earlypost-opAsepticPrevention ofn/aNot available,1-2 days or untilAntibiotic bonerevisioninfection arisingor itsthe surgical sitecement, bonefrom eitherreliabilityis open throughsubstitutes,contamination oris lowdrainage. Longerfreehand use ofa low-gradeif low-gradelocal antibioticinfectioninfection ispowder orsuspected.solutionSepticEradication of  100%AvailableSeveral weeksAntibiotic bonerevisionbacterialcement,infectionfreehand use oflocal antibioticpowder orsolution. Bonesubstitutes arecontraindicated.
It is evident that even if one focuses only on antibiotic bone substitutes several formulations should be available in order to meet these diverging criteria (Zilberman et al. J Control Release. 2008; 130:202-215). One way of modifying the release kinetics of drugs in an implantable formulation is to couple the active agent with biodegradable polymers. Two well known materials, which are frequently used to form biodegradable coatings are chitosan (Chi) and sodium alginate (Na-Alg) (Akter et al. Radiation Physics and Chemistry. 2012; 81:995-998, Buranapanitkit et al. Clin Orthop Relat Res. 2005:236-241). These bio-polymers have been investigated over a wide scale including the preparation of fibers, nanoparticles and even bone substitutes, thus they pose a very low risk of toxicity (Dai et al. J Biomed Biotechnol. 2009; 2009:595126, Sanna et al. Int J Nanomedicine. 2012; 7:5501-5516, Zhou et al. Int J Nanomedicine. 2013; 8:877-887). Chitosan is prepared from shrimp-shell chitin with hydrolysis and is only soluble in acidic media. It forms excellent films and coatings and in case it is added to acidic forms of drugs it can slow down release and degradation as described in WO 2009/050527 A1. 2009. Alginate derivatives such as alginic acid or sodium alginate are produced from seaweed species. The main feature of sodium alginate is that it is insoluble in acidic solutions and forms a biodegradable film that can be turned into water insoluble calcium alginate (Ca-Alg), which can act as a barrier for drug coatings. The general view of the surgical community is that local use of antibiotics without any carrier is only effective for the first few days post-op, however, this view is not supported by reliable experimental data. Theoretically it can be hypothesized that fixation of the antibiotic with physico-chemical means such as freeze-drying or embedding in polymer coatings may prolong the release of drugs, however, it is unknown if these procedures can meet the requirements detailed in Table 1.
The market of bone fillings and bone substitutes consist of a wide range of materials and biologically active components respectively.
As bone substitute materials, we can mention examples like tricalcium phosphate, calcium sulfate, hydroxyapatite and human bone grafts. The main functions of the substitute materials are mechanical strength, healing and structural re-integration of cells and tissues.
Biologically active coatings or pharmaceutically active agents include components (e.g. growth factors, antibiotics), which can enhance bone formation, cell adhesion, cell differentiation or can prevent bacterial infection.
More specifically defined bone substitutes include polymers that e.g. comprises of polylactic acid, polyglycolic acid or poly(lactic-co-glycolic acid). WO2012094708 discloses a synthetic bone replacement material that can prevent potential infection compared to human derived bone allograft. Also during human allograft disinfection prior to implantation, there is also a risk that the bone contains traces of disinfectant. However, Liban Chang et al. (Formosan Journal of Musculoskeletal Disorders 2 (2011) 55-61) showed with the use of supercritical CO2 a simple and safe method can be achieved to process human bone allograft disinfection.
Still till nowadays, bone is the most convenient grafting material, and according to the review article of Konstantinos Anagnostakos et al. (International Journal of Biomaterials, Volume 2012, Article ID 538061, 9 pages). This article describes a type of pharmaceutically active components, mainly focusing on antibiotics. The idea of mixing bone with antibiotics is known in the art, (De Grood et al. Ned Tijdschr Geneeskd, vol. 91.III.32, pp. 2192-2196, 1947) was the first to report on mixing penicillin with cancellous bone when filling bone defects in 1947. However emerging interest only appeared in the 1980s of mixing bone with biologically active components. The coating methods used were limited, manual mixing or incubation in solvents was the most frequently used techniques.
The used biologically active coatings mainly focus on antibiotics and cell adhesion and/or growth enhancers. As disclosed in the US Patent Application No. 20090324683 describing a biologically active coating (Bruce G Evans et al., “Controlled Release Tissue Graft Combination Biomaterials”) transforming growth factors (TGFs), bone morphogenetic proteins (BMPs), fibroblast growth factors (FGFs), parathyroid hormone derivatives (PTHs), Nell-1, statins, certain known osteoinductive peptides (e.g., P15, truncated PTHs or collagens), insulin-like growth factors (IGFs), and/or platelet-derived growth factors (PDGFs), or their respective therapeutic nucleotide transgenes may be used for this purpose.
The present invention comprises of tissue substitute material that is intended to be used for human implantation purposes, mainly to enhance cell adhesion and cell growth and to prevent bacterial infection. The used grafting material may be any such material customary in the field. The controlled release coating according to the invention may be used on any surface that enables the adsorption of the monomers/polymers utilized, or on any surface that may be made artificially adsorbent. In a preferred aspect, the grafting material is human tissue, preferably from the musculoskeletal system, more preferably cancellous bone allograft, which we found to be optimal for vascularisation and re-integration of existing tissues. The used antibiotic agents were selected by comparing practical results among these practical reasons are water solubility, heat stability and mechanical properties. The controlled release coating materials, which were chosen, are biocompatible, bioavailable. All the used materials used are already used in pharmacological products.
Accordingly, the present invention provides a tissue substitute material for implantation, comprising (a) a substrate made of the tissue to be implanted covered with (b) a controlled release coating containing (c) at least one biologically active substance that decreases bacterial growth, wherein the (b) controlled release coating is a bioavailable, biocompatible polymer material selected from the group consisting of: chitosan, alginic acid or a combination thereof, or any one of those in combination with pectin; and wherein the (c) at least one biologically active substance that decreases bacterial growth can be any antibiotics or a mixture thereof.
For the present invention, US 20090324683 could be considered as the closest prior art, since it relates to the same technical field and provides solutions similar in their scope. In view of this, to avoid unnecessary inflation of the extent of the present specification, Section A thereof with the definitions is included herein by reference.
US 20090324683 discloses tissue graft combination biomaterials comprising one or more agents, including bioactive agents, pharmaceutically active agents, or combinations thereof, which can be combination biomaterials of one or more agents and one or more substrates suitable for use as tissue graft materials. The disclosed combination tissue graft biomaterial comprises a biocompatible substrate; a degradable natural or synthetic polymer coated over the substrate surface; and a bioactive agent or pharmaceutically active agent encapsulated by the polymer matrix. By “encapsulated” is meant that the agent(s) can be either incorporated into the polymer or into or onto the substrate and covered by the polymer coating, such that release of the agent(s) from the combination tissue graft biomaterial is hindered and controlled by the polymer coating barrier and its degradation at the site of application. Also disclosed are methods of making the disclosed tissue grafts to select the rate of controlled release of bioactive agents, pharmaceutically active agents, or combinations thereof to produce therapy at the implant site.
First and foremost it should be noted that US 20090324683 describes a general, conceptualized controlled release system. This disclosure is expanded with laundry list type sections on all of the support component, the polymer component and the active agent. Contrary to the countless of the possible combinations of these three main constituent of the system, US 20090324683 gives experimental results only for polycaprolactone (PCL) as the controlled release polymer coating material, and a couple of antibiotics. It is evident that the controlled release properties of such a tripartite system are more dependent on the polymer component than either of the support or the active agent. In this respect, US 20090324683 clearly lacks enabling disclosure for the polymers in general.
More significantly, the example provided is for PCL, which is a water-insoluble material, the monomer of which is applied in acetone mixed with the active ingredients. It is apparent that such method is very limited in its application, may work with the antibiotics suggested in the experimental part of US 20090324683, but clearly not a good solution for more sensitive biomolecules, such as growth factors, hormones, etc. In summary, PCL coating can either be constructed using an organic solvent or heating the polymer to at least 50° C., which circumstances decrease biocompatibility and increase decomposition of the incorporated biologically active substance.
Further, there is not a single mention in US 20090324683 for the use of combination of polymers. It is clear, as detailed below that the combination of different polymers with pectin provides significantly improved properties for the controlled release coating. No such advantages could be foreseen based on the prior art.
Accordingly, to highlight the novelty and inventiveness of the present invention over the prior art, it is once again emphasized that the invention provides a bone substitute material for implantation, comprising the bone allograft to be implanted covered with a controlled release coating made of a polymer wherein the monomers of said polymer are water-soluble, and containing at least one biologically active substance that decreases bacterial growth. The polymer materials used form a valid selection over the prior art.
In addition, the methods of preparation of such a bone substitute material according to the invention are also clearly novel and inventive over the prior art as the use of organic phase may be completely eliminated during the manufacturing process. Further, the inventive production process uses a two-stage polymerization, which is made possible by the physic-chemical properties of the polymers used. Both chitosan and alginate that forms the base of the controlled release coating only form the polymer from the monomers where certain chemical changes induce the polymerization. The polymers used in the present invention are constructed from water soluble starting materials, all of them are low molecular weight substances (3-25 kDa).
In another aspect, the present invention provides a method for preparing a tissue substitute material for implantation, comprising
(a) preparing a homogenous coating on the substrate to be implanted from at least one biologically active substance that decreases bacterial growth;
(b) preparing a film coating from the water-soluble monomers of a biocompatible polymer material selected from the group consisting of chitosan and alginic acid, or a combination thereof, or any one of those in combination with pectin;
(c) drying the water-soluble film coating;
(d) converting the water soluble film coating into water insoluble film coating;
(e) drying the water-insoluble film coating.
In a preferred embodiment, the invention provides a tissue substitute material or method wherein the tissue is a tissue from the musculoskeletal system, preferably bone tissue, cartilage tissue or tendon tissue.
In another preferred embodiment, the invention provides a tissue substitute material or method wherein the tissue is bone allograft.
In a particularly preferred embodiment, the invention provides a tissue substitute material for implantation which is a bone substitute material for implantation, comprising (a) the bone allograft to be implanted covered with (b) a controlled release coating containing (c) at least one biologically active substance that decreases bacterial growth, wherein the (b) controlled release coating is a bioavailable, biocompatible polymer material selected from the group consisting of: chitosan, alginic acid or a combination thereof, or any one of those in combination with pectin; and wherein the (c) at least one biologically active substance that decreases bacterial growth can be any antibiotics or a mixture thereof.
In another particularly preferred embodiment, the invention provides a method for preparing a tissue substitute material for implantation, which is a bone allograft material for implantation, comprising
(a) preparing a homogenous coating on the bone allograft material from at least one biologically active substance that decreases bacterial growth;
(b) preparing a film coating from the water-soluble monomers of a biocompatible polymer material selected from the group consisting of chitosan and alginic acid, or a combination thereof, or any one of those in combination with pectin;
(c) drying the water-soluble film coating;
(d) converting the water soluble film coating into water insoluble film coating;
(e) drying the water-insoluble film coating.
In another embodiment, the invention provides a tissue substitute material or method, wherein the substrate is a known tissue substitute, preferably bone substitute, more preferably selected from the group consisting of implants made from metal, plastic or other materials, and standalone polymer material suitable for the preparation of the coating, preferably alginate beads.
In this specific embodiment of the invention, a particularly preferred substrate is the standalone polymer material that otherwise also used for the preparation of the coating. More particularly, the combination of pectin and sodium alginate results in a unique polymer combination, which can be used not only as coating, but as solid microspheres.
The art discloses several substrate systems which are formed from this kind of polymer materials. However, pectin is not used as a gelling agent or thickener according to the present invention, but as a water soluble biodegradable part of a two or more component biocompatible polymer system, which is partly water insoluble. The inventors surprisingly found that the water soluble Na-alginate part of the complex polymer can be selectively converted into water insoluble Ca-alginate but leaves the water soluble pectin part intact with choosing the appropriate Ca2+ source and the time of Na—Ca conversion. The film coating or microsphere prepared this way enables pectin to elute freely in the aqueous media depending on the concentration and distribution in the polymer system, while Ca-alginate mainly releases the drug during decomposition. Thus the release of the incorporated drug content can increase or decrease depending on the pectin-alginate (as water soluble-water insoluble) ratio.
In another preferred embodiment, the invention provides a tissue substitute material wherein the (b) controlled release coating is alginic acid pectin copolymer.
In a further preferred embodiment, the invention provides a tissue substitute material or method wherein the (b) controlled release coating essentially consists of alginic acid within the range of 70 to 90% and pectin within the range of 10 to 30%.
In another preferred embodiment, the invention provides a tissue substitute material or method wherein the (c) at least one biologically active substance that decreases bacterial growth is selected from the group consisting of gentamicin, ciprofloxacin, vancomycin, amoxicillin.
In another preferred embodiment, the invention provides a tissue substitute material or method, further comprising other biologically active ingredients to enhance cell migration, adhesion and growth.
In particularly preferred embodiments, the invention provides a tissue substitute material or method according said biologically active ingredient to enhance cell migration, adhesion and growth is selected from the group consisting of growth factors including PDGF, TGF-/315 vascular endothelial growth factor, basic fibroblast growth factor (bFGF), and epidermal growth factor; albumin, platelet rich plasma (PRP), platelet pure plasma (PPP) and platelet rich fibrin (PRF) and other blood separation products that contain cell growth and/or cell migration enhancing agents.
In a preferred embodiment of the method of the invention, the antibiotic coating is prepared in step (a) by freeze drying, solvent evaporation or vacuum evaporation. In a preferred embodiment of the method of the invention, the film coating is prepared in step (a) spraying or casting.
In another preferred embodiment of the method of the invention, the drying in step (c) and/or (e) is accomplished in a drying chamber or exsiccator, or by using moderate heating and vacuum. In a further preferred embodiment of the method of the invention, in step (d), the conversion of the soluble film coating into water insoluble film coating is accomplished by using a Ca2+ ion containing solution, preferably within the range of 1 to 20%.
In another aspect, the invention provides the tissue substitute material, preferably a bone allograft material for implantation according to the invention or the bone allograft material obtainable by the method according to the invention, for the treatment of a condition, disease or disorder selected from the group consisting of:
i, prevention of bone infection. e.g. Surgery, contaminated wounds, open fractures, implantation of any foreign material, presence of any foreign material, filling the cavity of bone cysts, treatment of aseptic non-union;
ii, prevention of re-infection. e.g. Revision surgery after septic complications, reconstructive surgery after traumatic or other bone loss, treatment of posttratumatic or post-septic non-union;
iii, treatment of bone infection. e.g. Acute or chronic osteomyelitis, ostitis, septic non-union, septic implants or prosthetic devices or any other foreign material, including projectiles.
In another aspect, the invention provides a method for treating a condition, disease or disorder selected from the group consisting of:
i, prevention of bone infection. e.g. Surgery, contaminated wounds, open fractures, implantation of any foreign material, presence of any foreign material, filling the cavity of bone cysts, treatment of aseptic non-union;
ii, prevention of re-infection. e.g. Revision surgery after septic complications, reconstructive surgery after traumatic or other bone loss, treatment of posttratumatic or post-septic non-union;
iii, treatment of bone infection. e.g. Acute or chronic osteomyelitis, ostitis, septic non-union, septic implants or prosthetic devices or any other foreign material, including projectiles,
wherein said method comprising the step implanting the bone allograft material for implantation according to the invention or the bone allograft material obtainable by the method according to the invention.